Abstract
Argonaute (Ago) proteins mediate silencing of nucleic acid targets by small RNAs. In fission yeast, Ago1, Tas3 and Chp1 assemble into a RITS complex, which silences transcription near centromeres. Here we describe a repetitive motif within Tas3, termed the 'Argonaute hook', that is conserved from yeast to humans and binds Ago proteins through their PIWI domains in vitro and in vivo. Site-directed mutation of key residues in the motif disrupts Ago binding and heterochromatic silencing in vivo. Unexpectedly, a PIWI domain pocket that binds the 5' end of the short interfering RNA guide strand is required for direct binding of the Ago hook. Moreover, wild-type but not mutant Ago hook peptides derepress microRNA-mediated translational silencing of a target messenger RNA. Proteins containing the conserved Ago hook may thus be important regulatory components of effector complexes in RNA interference.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Amino Acid Sequence*
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Argonaute Proteins
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Carrier Proteins* / genetics
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Carrier Proteins* / metabolism
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Cell Cycle Proteins / genetics
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Cell Cycle Proteins / metabolism
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Cell Nucleus / metabolism
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Eukaryotic Initiation Factor-2 / genetics
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Eukaryotic Initiation Factor-2 / metabolism
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Gene Silencing
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Humans
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Molecular Sequence Data
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Mutagenesis, Site-Directed
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Protein Binding
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Protein Biosynthesis
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Protein Conformation*
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RNA Interference
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RNA, Small Interfering / genetics
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RNA, Small Interfering / metabolism
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RNA-Binding Proteins
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Schizosaccharomyces / genetics
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Schizosaccharomyces / metabolism
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Schizosaccharomyces pombe Proteins* / chemistry
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Schizosaccharomyces pombe Proteins* / genetics
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Schizosaccharomyces pombe Proteins* / metabolism
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Sequence Alignment
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Two-Hybrid System Techniques
Substances
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AGO2 protein, human
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Ago1 protein, S pombe
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Argonaute Proteins
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Carrier Proteins
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Cell Cycle Proteins
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Chp1 protein, S pombe
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Eukaryotic Initiation Factor-2
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RNA, Small Interfering
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RNA-Binding Proteins
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Schizosaccharomyces pombe Proteins
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Tas3 protein, S pombe